Microbial Identification

1MICROBIAL IDENTIFICATION USING THE BIOMÉRIEUX VITEK® 2 SYSTEM David H. Pincus bioMérieux, Inc. Hazelwood, MO, USA OBJECTIVE This chapter describes the VITEK 2 automated microbiology system and its application in the identification of microorganisms. PRINCIPLES The VITEK 2 is an automated microbiology system utilizing growth-based technology. The system is available in three formats (VITEK 2 compact, VITEK 2, and VITEK 2 XL) that differ in increasing levels of capacity and automation. Figure 1 shows the VITEK 2 compact system. All three systems accommodate the same colorimetric reagent cards that are incubated and interpreted automatically. 1 www.pda.org/bookstore 2 Encyclopedia of Rapid Microbiological Methods Figure 1. VITEK 2 Compact Instrument and Workstation. VITEK 2 Compact This format focuses on the industrial microbiology-testing environment while also having application for low to middle volume clinical laboratories. Features specifically developed for industrial microbiology include 21 CFR Part 11 compliance (for electronic records and signatures) and a colorimetric reagent card (BCL) used to identify the spore-forming Gram-positive bacilli (i.e., Bacillus and related genera). The other colorimetric reagent cards (GN, GP, YST) apply to all system formats for both industrial and clinical laboratories. VITEK 2 and VITEK 2 XL These formats are more focused on the clinical microbiology laboratory and provide increased levels of automation and capacity for higher volume laboratories. They also provide an option of automatic pipetting and dilution for antimicrobial susceptibility testing. Reagent Cards The reagent cards have 64 wells that can each contain an individual test substrate. Substrates measure various metabolic activities such as acidification, alkalinization, enzyme hydrolysis, and growth in the presence of inhibitory substances. An optically clear film present on both sides of the card allows for the appropriate level of oxygen transmission while maintaining a sealed vessel www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 3 that prevents contact with the organism-substrate admixtures. Each card has a pre-inserted transfer tube used for inoculation (described below). Cards have bar codes that contain information on product type, lot number, expiration date, and a unique identifier that can be linked to the sample either before or after loading the card onto the system. Figure 2 shows the GN card. Figure 2. VITEK 2 GN Colorimetric Identification Card. There are currently four reagent cards available for the identification of different organism classes as follows: 1. GN - Gram-negative fermenting and non-fermenting bacilli 2. GP - Gram-positive cocci and non-spore-forming bacilli 3. YST - yeasts and yeast-like organisms 4. BCL - Gram-positive spore-forming bacilli Product-specific details for each of the identification cards are shown below. Culture Requirements The on-line product information contains a culture requirements table that lists parameters for appropriate culture and inoculum preparation. These parameters include acceptable culture media, culture age, incubation conditions, and inoculum turbidity. www.pda.org/bookstore 4 Encyclopedia of Rapid Microbiological Methods Suspension Preparation A sterile swab or applicator stick is used to transfer a sufficient number of colonies of a pure culture and to suspend the microorganism in 3.0 mL of sterile saline (aqueous 0.45% to 0.50% NaCl, pH 4.5 to 7.0) in a 12 x 75 mm clear plastic (polystyrene) test tube. The turbidity is adjusted accordingly (see Table 1) and measured using a turbidity meter called the DensiChekTM. Table 1: Suspension Turbidities Used for Card Inoculation. �� Inoculation Identification cards are inoculated with microorganism suspensions using an integrated vacuum apparatus. A test tube containing the microorganism suspension is placed into a special rack (cassette) and the identification card is placed in the neighboring slot while inserting the transfer tube into the corresponding suspension tube. The cassette can accommodate up to 10 tests (VITEK 2 Compact; see Figure 3) or up to 15 tests (VITEK 2 and VITEK 2 XL; see Figure 4). The filled cassette is placed either manually (VITEK 2 compact) or transported automatically (VITEK 2 and VITEK 2 XL) into a vacuum chamber station. After the vacuum is applied and air is re-introduced into the station, the organism suspension is forced through the transfer tube into micro-channels that fill all the test wells. Card Sealing and Incubation Inoculated cards are passed by a mechanism, which cuts off the transfer tube and seals the card prior to loading into the carousel incubator. The carousel incubator can accommodate up to 30 or up to 60 cards. All card types are www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 5 incubated on-line at 35.5 + 1.0ºC. Each card is removed from the carousel incubator once every 15 minutes, transported to the optical system for reaction readings, and then returned to the incubator until the next read time. Data are collected at 15-minute intervals during the entire incubation period. Figure 3. VITEK 2 Compact Cassette Loaded with 10 Cards and Suspension Tubes and Bar Code Scanner for Data Entry. Figure 4. VITEK 2 Cassette Loaded with Cards and Suspension Tubes Being Loaded Into the Automatic Transport System. www.pda.org/bookstore 6 Encyclopedia of Rapid Microbiological Methods Optical System A transmittance optical system allows interpretation of test reactions using different wavelengths in the visible spectrum. During incubation, each test reaction is read every 15 minutes to measure either turbidity or colored products of substrate metabolism. In addition, a special algorithm is used to eliminate false readings due to small bubbles that may be present. Test Reactions Calculations are performed on raw data and compared to thresholds to determine reactions for each test. On the VITEK 2 Compact, test reaction results appear as “+”,” –“, “(–)” or “(+)”. Reactions that appear in parentheses are indicative of weak reactions that are too close to the test threshold. With the VITEK 2 or VITEK 2 XL, these weak reactions appear as “?”. Database Development The databases of the VITEK 2 identification products are constructed with large strain sets of well-characterized microorganisms tested under various culture conditions. These strains are derived from a variety of clinical and industrial sources as well as from public (e.g., ATCC) and university culture collections. Analytical Techniques Test data from an unknown organism are compared to the respective database to determine a quantitative value for proximity to each of the database taxa. Each of the composite values is compared to the others to determine if the data are sufficiently unique or close to one or more of the other database taxa. If a unique identification pattern is not recognized, a list of possible organisms is given, or the strain is determined to be outside the scope of the database. Identification Levels An unknown biopattern is compared to the database of reactions for each taxon, and a numerical probability calculation is performed. Various qualitative levels of identification are assigned based on the numerical probability calculation. The different levels and associated information are shown in Table 2. www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 7 Mixed Taxa Identifications This occurs when the biopattern is representative of a collective taxon and generates a genus-level, group-level, or slashline identification. In rare cases, species-level identification can be a mixed taxon comprised of two subspecies. Supplemental tests may be used to delineate representative species or subspecies of these collective taxa. Supplemental Testing In the case of low discrimination identifications, two or three choices are listed in the order of their probability calculations. However, all taxa appearing in a low discrimination identification are viable choices and should only be ruled out after additional testing and/or observation. The lab report contains recommended supplemental tests that allow for differentiation of choices in low discrimination identifications. If the supplemental tests listed are insufficient to complete the identification, then standard microbiology references should be consulted. Table 2. Identification Levels. �� www.pda.org/bookstore 8 Encyclopedia of Rapid Microbiological Methods Non-Reactive Biopattern When a biopattern is calculated for an unknown organism that is either completely negative or consists of both negative tests and tests with reactions that lie too close to the test thresholds, the identification result will be “Nonreactive biopattern.” If one encounters a non-reactive biopattern, a note will appear that states: “Organism with low reactivity biopattern – please check viability.” The species that can potentially trigger this note are shown in separate tables for each of the respective products described below. APPLICATIONS GN Card The GN card is used for the automated identification of 135 taxa of the most significant fermenting and non-fermenting Gram-negative bacilli. The list of claimed species is shown in Table 3. Of the 135 taxa, 12 are grouped taxa in either genus (3), group (6), or slashline (3) designations. When representative taxa of these collective designations are included, the total number of taxa claimed by GN is over 160. As with low discrimination identifications, grouped taxa can be separated into their component taxa by supplemental testing and/or observation. A list of the grouped taxa is shown in Table 4. There are informational notes that appear with certain taxa and these are shown in Table 5. When no reactivity is encountered, one is referred to a list of species that may be associated with a non-reactive biopattern (see Table 6). The GN card is based on established biochemical methods and newly developed substrates measuring carbon source utilization, enzymatic activities, and resistance (ASM 1998, Atlas 1993, Brenner et al. 1993, Chang et al. 2002, Coenye et al. 2001a, Coenye et al. 2001b, De Baere et al. 2001, Freney et al. 2000, Gavini et al. 1989, Holt et al. 1994, Krieg and Holt 1984, Murray et al. 1999, Richard and Kiredjian 1992, Smith et al. 1991, Vandamme et al. 1999). There are 47 biochemical tests and one negative control well. Final identification results are available in approximately 10 hours or less. The list of test substrates is shown in Table 7. In a recent multi-site study, the performance of the VITEK 2 GN was evaluated using 562 isolates of both commonly and rarely observed species of www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 9 Gram-negative bacilli, including 153 non-fermentative strains. The reference identification was determined with api® 20 E and api 20 NE identification kits. Overall, the VITEK 2 GN correctly identified 96.8% of the isolates, including 6.4% low discrimination with the correct species listed. Misidentifications occurred at 3.0% and no identifications occurred at 0.2%. GP Card The GP card is used for the automated identification of 115 taxa of the most significant non-spore-forming Gram-positive bacteria (primarily cocci). The list of claimed species is shown in Table 8. Of the 115 taxa, seven are grouped taxa in either slashline (4) or species (3) designations. When representative species or subspecies of these collective designations are included, the total number of taxa claimed by GP is over 120. As with low discrimination identifications, grouped taxa can be separated into their component taxa by supplemental testing and/or observation. A list of the grouped taxa is shown in Table 9. There are informational notes that appear with certain taxa and these are shown in Table 10. When no reactivity is encountered, one is referred to a list of species that may be associated with a non-reactive biopattern (see Table 11). The GP identification card is based on established biochemical methods and newly developed substrates (Atlas 1993, Barros et al. 2001, Bille et al. 1992, Collins et al. 1984a, Collins et al. 1984b, Collins and Lawson 2000, Collins et al. 2001, Coykendall 1989, Devriese et al. 1988, Farrow et al. 1989, Freney et al. 2000, Holt et al. 1994, Kilpper-Bälz and Schleifer 1987, Krieg and Holt 1984, Murray et al. 1999, Poyart et al. 2002, Schlegel et al. 2000, Viera et al. 1998, Whiley et al. 1999). There are 43 biochemical tests measuring carbon source utilization, enzymatic activities and resistance. Final identification results are available in approximately eight hours or less. The list of test substrates is shown in Table 12. In a recent multi-site study, the performance of the VITEK 2 GP was evaluated using 457 isolates of both commonly and rarely observed species of Gram-positive cocci. The reference identification was determined with api STAPH and api 20 STREP identification kits. Overall, the VITEK 2 GP correctly identified 96.5% of the isolates, including 2.2% low discrimination with the correct species listed. Misidentifications occurred at 3.3% and no identifications occurred at 0.2%. www.pda.org/bookstore 10 Encyclopedia of Rapid Microbiological Methods Table 3. Organisms Identified by the GN Card. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 11 Table 3. Organisms Identified by the GN Card (continued). �� www.pda.org/bookstore 12 Encyclopedia of Rapid Microbiological Methods Table 4. GN Grouped Taxa. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 13 Table 5. Notes Associated with Certain GN Taxa. �� Table 6. Species that May Be Non-reactive on the GN Card. �� www.pda.org/bookstore 14 Encyclopedia of Rapid Microbiological Methods Table 7. Test Substrates on GN Card. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 15 Table 8. Organisms Identified by the GP Card. �� www.pda.org/bookstore 16 Encyclopedia of Rapid Microbiological Methods Table 8. Organisms Identified by the GP Card (continued). �� Table 9. GP Grouped Taxa. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 17 Table 10. Notes Associated with Certain GP Taxa. �� Table 11. Species That May Be Non-Reactive on the GP Card. �� www.pda.org/bookstore 18 Encyclopedia of Rapid Microbiological Methods Table 12. Test Substrates on GP Card. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 19 YST Card The YST card is used for the automated identification of 49 taxa of the most significant yeasts and yeast-like organisms. The list of claimed species is shown in Table 13. Of the 49 taxa, three are grouped taxa in either slashline (2) or genus (1) designations. When representative species of these collective designations are included, the total number of taxa claimed by YST is 54. As with low discrimination identifications, grouped taxa can be separated into their component taxa by supplemental testing and/or observation. A list of the grouped taxa is shown in Table 14. When no reactivity is encountered, one is referred to a list of species that may be associated with a non-reactive biopattern (see Table 15). The YST identification card is based on established biochemical methods and newly developed substrates (Atlas 1993, Barnett et al. 2000, Kreger-van Rij 1984, Larone 1995, Lodder 1971, McGinnis 1980, Murray et al. 1999). There are 46 biochemical tests measuring carbon source utilization, enzymatic activities and resistance. Final identification results are available in approximately 18 hours. The list of test substrates is shown in Table 16. In a recent multi-site study, the performance of the VITEK 2 YST was evaluated using 623 isolates of both commonly and rarely observed species of yeast and yeast-like organisms. The reference identification was determined with api 20C AUX identification kits. Overall, the VITEK 2 YST correctly identified 98.9% of the isolates, including 11.7% low discrimination with the correct species listed. Misidentifications occurred at 0.6% and no identifications occurred at 0.5%. www.pda.org/bookstore 20 Encyclopedia of Rapid Microbiological Methods Table 13. Organisms Identified by the YST Card. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 21 Table 14. YST Grouped Taxa. �� Table 15. Species That May Be Non-Reactive on the YST Card. �� www.pda.org/bookstore 22 Encyclopedia of Rapid Microbiological Methods Table 16. Test Substrates on YST Card. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 23 BCL Card The BCL card is used for the automated identification of 38 taxa of the most significant aerobic endospore-forming species of the family Bacillaceae. The list of claimed species is shown in Table 17. Of the 38 taxa, four are grouped taxa in slashline designations. When representative species of these slashlines are included, the total number of taxa claimed by BCL is 42. As with low discrimination identifications, grouped taxa can be separated into their component taxa by supplemental testing and/or observation. A list of the slashline taxa is shown in Table 18. There are informational notes that appear with certain taxa and these are shown in Table 19. When no reactivity is encountered, one is referred to a list of species that may be associated with a non-reactive biopattern (see Table 20). The BCL identification card is based on established biochemical methods and newly developed substrates (Atlas 1993, Claus and Berkeley 1986, Gordon et al. 1973, Logan and Berkeley 1981, Logan and Berkeley 1984, Logan et al. 1985, Logan et al. 2002, Logan and Turnbull 2003). There are 46 biochemical tests measuring carbon source utilization, enzymatic activities, inhibition, and resistance. Final identification results are available in approximately 14 hours. The list of test substrates is shown in Table 21. The database performance of the VITEK 2 BCL was evaluated using 1436 isolates of both commonly and rarely observed species of Gram-positive aerobic spore-forming bacilli. The reference identification was determined with the api 50CHB identification kit and other conventional test methods. Overall, the VITEK 2 BCL correctly identified 96.1% of the isolates, including 9.3% low discrimination with the correct species listed. Misidentifications occurred at 2.6% and no identifications occurred at 1.3%. www.pda.org/bookstore 24 Encyclopedia of Rapid Microbiological Methods Table 17. Organisms Identified by the BCL Card. �� Table 18. BCL Slashline Taxa. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 25 Table 19. Notes Associated with Certain BCL Taxa. �� Table 20. Species That May Be Non-Reactive on the BCL Card. �� www.pda.org/bookstore 26 Encyclopedia of Rapid Microbiological Methods Table 21.Test Substrates on BCL Card. �� www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 27 VALIDATION PROCEDURE Quality control organisms and their expected results are listed for each product and are tested according to the procedures outlined in the on-line product information. Frequency of testing is dependent on local regulations. POTENTIAL NEW APPLICATIONS A new application to be released soon is the NH card, used to identify species of Neisseria, Haemophilus, and other fastidious genera including Campylobacter. CONCLUSION The VITEK 2 is an automated microbial identification system that provides highly accurate and reproducible results as shown in multiple independent studies. With its colorimetric reagent cards, and associated hardware and software advances, the VITEK 2 offers a state-of-the-art technology platform for phenotypic identification methods. REFERENCES American Society for Microbiology. 98th General Meeting Workshop Program. Practical Approach to the Identification of the Medically Important Glucose Non-Fermenting Gram-Negative Bacilli. American Society for Microbiology, Washington, D.C. 1998. Atlas RA. Handbook of Microbiological Media. CRC Press, Ann Arbor. 1993. Barnett JA, Payne RW, Yarrow D, editors. Yeasts: Characteristics and Identification, 3rd ed. Cambridge University Press, New York. 2000. Barros RR, Carvalho GS, Peralta JM, Facklam RR, Teixeira LM. Phenotypic and Genotypic Characterization of Pediococcus Strains Isolated from Human Clinical Sources. J. Clin. Microbiol. 2001;39:1241-1246. www.pda.org/bookstore 28 Encyclopedia of Rapid Microbiological Methods Bille J, Catimel B, Bannerman E, Jacquet C, Yersin MN, Caniaux I, Monget D, Rocourt J. API Listeria, a New and Promising One-Day System to Identify Listeria Isolates. Appl. Environ. Microbiol. 1992;58:1857-1860. Brenner DJ, Grimont PAD, Steigerwalt AG, Fanning GR, Ageron E, Riddle CF. Classification of Citrobacteria by DNA Hybridization: Designation of Citrobacter farmeri sp.nov., Citrobacter youngae sp.nov., Citrobacter braakii sp.nov., Citrobacter werkmanii sp.nov., Citrobacter sedlakii sp.nov., and Three Unnamed Citrobacter Genomospecies. Int. J. Syst. Bacteriol. 1993; 43:645-658. Chang YH, Han J, Chun J, Lee KC, Rhee MS, Kim YB, Bae KS. Comamonas koreensis sp.nov., a non-motile species from wetland in Woopo, Korea. Int. J. Syst. Evol. Microbiol. 2002;52:377-318. Claus, D., & Berkeley, R. C. W. 1986. Genus Bacillus Cohn 1872. In Bergey’s Manual of Systematic Bacteriology, Vol. 2, pp. 1105-1139. Edited by P. H. A. Sneath, N. S. Mair, M. E. Sharpe & J. G. Holt. Baltimore: Williams & Wilkins. Coenye T, Mahenthiralingam E, Henry D, Lipuma JJ, Laevens S, Gillis M, Speert DP, Vandamme P. Burkholderia ambifaria sp nov., a novel member of the Burkholderia cepacia complex including biocontrol and cystic fibrosis-related isolates. Int. J. Syst. Evol. Microbiol. 2001;51:1481-1490. Coenye T, Vandamme P, Gowan JRW, Lipuma JJ. Taxonomy and Identification of the Burkholderia cepacia Complex. J. Clin. Microbiol. 2001;39:3427-3436. Collins MD, Farrow JAE, Katic V, Kandler O. Taxonomic studies on streptococci of serological groups E, P, U and V: description of Streptococcus porcinus sp. nov. Syst. Appl. Microbiol. 1984;5:402-413. Collins MD, Jones D, Farrow JAE, Kilpper-Bälz R, Schleifer KH. Enterococcus avium nom. rev., comb. nov.; E.casseliflavus nom. rev., comb. nov.; E. durans nom. rev., comb. nov.; E. gallinarum comb. nov.; and E. malodoratus sp.nov. Int. J. Syst. Bacteriol. 1984;34:220-223. Collins MD, Lawson PA. The genus Abiotrophia (Kawamura et al.) is not monophiletic: proposal of Granulicatella gen. nov., Granulicatella adiacens comb.nov., Granulicatella elegans comb. nov.and Granulicatella balaenopterae comb. nov. Int. J. Syst. Evol. Microbiol. 2000;50:365-369. www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 29 Collins MD, Hutson RA, Hoyles L, Falsen E, Nikolaitchouk N, Foster G. Streptococcus ovis sp. nov. isolated from sheep. Int. J. Syst. Evol. Microbiol. 2001;51:1147-1150. Coykendall AL. Classification and Identification of the Viridans Streptococci. Clin. Microbiol. Rev. 1989;2:315-328. De Baere T, Steyaert, Wauters G, De Vos P, Goris J, Coenye T, Suyama T, Verschraegen G, Vaneechoutte M. Classification of Ralstonia pickettii biovar 3/ ‘thomasii’ strains (Pickett 1994) and of new isolates related to nosocomial recurrent meningitis as Ralstonia mannitolytica sp.nov. Int. J. Syst. Evol. Microbiol. 2001;51:547-558. Devriese LA, Kilpper-Bälz R, Schleifer KH. Streptococcus hyointestinalis sp.nov. from the gut of swine. Int. J. Syst. Bacteriol. 1988;38:440-441. Farrow JAE, Facklam RR, Collins MD. Nucleic acid homologies of some vancomycin-resistant leuconostocs and description of Leuconostoc citreum sp. nov. and Leuconostoc pseudomesenteroides sp. nov. Int. J. Syst. Bacteriol. 1989;39:279-283. Freney J, Renaud F, Hansen W, Bollet C. Précis de bactériologie clinique, ESKA, Paris, France. 2000. Gavini F, Mergaert J, Beji A, Mielcarek C, Izard D, Kersters K, DeLey J. Transfer of Enterobacter agglomerans (Beijerinck 1888) Ewing and Fife to Pantoea gen. nov. as Pantoea agglomerans comb.nov. and Description of Pantoea dispersa sp. nov. Int. J. Syst. Bacteriol. 1989;39:337-345. Gordon, R. E., Haynes, W. C., Pang, C. H. N. 1973. The genus Bacillus. In Agriculture handbook no. 427, pp. 283. U.S. Department of Agriculture. Washington, D.C. Holt JG, Krieg NR, Sneath PHA, Staley JT, Williams ST. Bergey’s Manual of Determinative Bacteriology, 9th ed. Williams and Wilkins, Baltimore, Maryland. 1994. Kilpper-Bälz R, Schleifer KH. Streptococcus suis sp. nov., nom. rev. Int. J. Syst. Bacteriol. 1987;37:160-162. www.pda.org/bookstore 30 Encyclopedia of Rapid Microbiological Methods Kreger-van Rij NJW, editor. The yeasts—a taxonomic study, 3rd ed. Elsevier Science Publishers B.V. Amsterdam. 1984. Krieg NR, Holt JG. Bergey’s Manual of Systematic Bacteriology, volume 1. William & Wilkins, Baltimore, Maryland. 1984. Larone DH. Medically Important Fungi—a guide to identification. 3rd ed. ASM Press. American Society for Microbiology. Washington, D.C. 1995. Lodder J. The Yeasts, 2nd ed. North Holland Publishing Company, Netherlands. 1971. Logan, N.A., & Berkeley, R.C.W. 1981. Classification and identification of members of the genus Bacillus. In The Aerobic Endospore-forming Bacteria, pp. 105-140. Edited by R.C.W. Berkeley & M. Goodfellow. Academic Press, London. Logan, N. A. & Berkeley, R. C. W. 1984. Identification of Bacillus strains using the API system. J. Gen. Microbiol. 130: 1871. Logan, N. A., Carman, J. A., Melling, J., Berkeley, R. C. W. 1985. Identification of Bacillus anthracis by API tests. J. Med. Microbiol. 20: 75. Logan, N. A., Forsyth, G., Lebbe, L., Goris, L., Heyndrickx, M., Balcaen, A., Verhelst, A., Falsen, E., Ljungh, Å., Hansson, H. B., DeVos, P. 2002. Polyphasic identification of Bacillus and Brevibacillus strains from clinical, dairy and industrial specimens and proposal of Brevibacillus invocatus sp. nov. Int. J. Syst. Evol. Microbiol. 52:953. Logan, N. A., Turnbull, P. C. B. 2003. Bacillus and recently derived genera. In Manual of Clinical Microbiology, 8th ed., pp. 445-460. Edited by P. R. Murray, E. J. Baron, M. A. Pfaller, J. H. Jorgensen & R. H. Yolken. American Society for Microbiology, Washington, DC. McGinnis MR. Laboratory Handbook of Medical Mycology, Academic Press, New York. 1980. Murray PR, Baron EJ, Pfaller MA, Tenover FC, Yolken RH, editors. Manual Of Clinical Microbiology, 7th ed. American Society for Microbiology, Washington, D.C. 1999. www.pda.org/bookstore Microbial Identification Using the bioMérieux VITEK® . . . 31 Poyart C, Quesne G, Trieu-Cuot P. Taxonomic dissection of the Streptococcus bovis group by analysis of manganese-dependent superoxide dismutase gene (sodA) sequences: reclassification of Streptococcus infantarius subsp. coli as Streptococcus lutetiensis sp.nov. and of Streptococcus bovis biotype II.2 as Streptococcus pasteurianus sp nov. Int. J. Syst. Evol. Microbiol. 2002;52:12471255. Richard C, Kiredjian M. Laboratory methods for the Identification of the Medically Important Glucose Nonfermenting Gram-Negative Bacilli. Institut Pasteur, Paris, France. 1992. Schlegel L, Grimont F, Collins MD, Regnault B, Grimont PAD, Bouvet A. Streptococcus infantarius sp. nov., Streptococcus infantarius subsp infantarius subsp. nov. and Streptococcus infantarius subsp coli subsp. nov., isolated from humans and food. Int. J. Syst. Evol. Microbiol. 2000;50:1425-1434. Smith SK, Sutton DC, Fuerst JA, Reichelt JL. Evaluation of the Genus Listonella and the reassignment of Listonella damsela (Love et al.) MacDonell and Colwell to the Genus Photobacterium as Photobacterium damsela comb. nov. with an Emended Description. Int. J. Syst. Bacteriol. 1991;41:529-534. Vandamme P, Goris J, Coenye T, Hoste B, Janssens D, Kersters K, DeVos P, Falsen E. Assignment of Centers for Disease Control group IVc-2 to the genus Ralstonia as Ralstonia paucula sp.nov. Int. J. Syst. Bacteriol. 1999;49:663-669. Viera VV, Teixeira LM, Zahner V, Momen H, Facklam RR, Steigerwalt AG, Brenner DJ, Castro ACD. Genetic relationships among the different phenotypes of Streptococcus dysgalactiae strains. Int. J. Syst. Bacteriol. 1998;48:1231-1243. Whiley RA, Hall LMC, Hardie JM, Beighton D. A study of small colony beta hemolytic, Lancefield group C streptococci within the anginosus group: description of Streptococcus constellatus subsp. pharynges subsp.nov., associated with the human throat and pharyngitis. Int. J. Syst. Bacteriol. 1999;49:1443-1449. www.pda.org/bookstore 32 Encyclopedia of Rapid Microbiological Methods ABOUT THE AUTHOR David H. Pincus, M.S., is Director of R&D Microbiology at bioMérieux’s St. Louis, USA facility. His background is medical microbiology with expertise in phenotypic identification product development and medical mycology. He was a bacteriologist in a clinical lab for 2 years before joining bioMérieux in 1978 as a mycologist for Analytab (“API-USA”). He acquired extensive experience in the reference laboratories in microbial identification/taxonomy with specialty in mycology before moving into R&D in 1988. He is currently focused on reagent development/support for VITEK/VITEK 2 systems and serves on the Editorial Boards of the Journal of Clinical Microbiology and Medical Mycology. www.pda.org/bookstore

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